In known bio-optical sensors, the reaction takes place on a surface of the semiconductor device which is an image surface divided into pixels. The light produced by reactions of this nature is small, and accordingly, the signal produced by any pixel of the device is also small. The signal is frequently less than other effects such as dark current (leakage current) from the pixel and voltage offsets. Therefore, a calibration/cancellation scheme may be necessary to increase the sensitivity of the system.
In the related field of solid state image sensors, there are a number of known calibration techniques. In image sensors, it may be necessary to have a continuous image plane on which the image is formed. Calibration techniques involve either the use of dark frame cancellation or the use of special calibration pixels.
In dark frame cancellation, a dark reference frame is taken and the resulting signal output is subtracted from the image frame. The dark reference frame is usually taken with the same exposure. The integration time is the same as the image but no light is impinged on the sensor, either by use of a shutter or by turning off the scene illumination.
When calibration pixels are used, they are provided at the edge of the sensor. The calibration pixels are usually in the form of a single row or column, since it is necessary to have a continuous image surface.
In current bio-optical sensors, a dark image is acquired before the analyte and reagents are deposited on the sensor. This calibration image is used during detection and processing of the photo-signal. This means that there is a time difference between the acquisition of the dark reference frame and the detection and processing of the sensor signal. During this time there may be changes in the conditions on the device, e.g., operating voltage and temperature may change due to a low battery, a change in the ambient temperature, or self-heating due to power dissipation. Consequently, the calibration signal is not an accurate representation of the dark signal at the relevant time.